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低温
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上海圻明生物
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2μg
折光马尔太虫PCR阳性对照质粒上海圻明生物优势供应。更多产品资料欢迎免费咨询。
One of the many important uses of PCR technology is that it can be used to label DNA probes with high specific activity. PCR technology has high specificity, and can synthesize probe DNA fragments in quantities within 1~2h if [α-32P]dNTP or other markers are added to the substrate
dNTPs, the probe DNA can be well labeled during DNA synthesis, and the incorporation rate of the marker can be as high as 70%~80%. Therefore, PCR labeling technology is particularly suitable for large-scale detection and non-radiolabeling. The disadvantage of this method is that a specific pair of PCR primers is synthesized.
Labeling can also be achieved by using small fragments prepared from probe DNA as primers.
Solution preparation
1. Prepare a stock solution
Unless otherwise stated, all unused stock solutions should be divided into disposable aliquots and stored at -20 °C after preparation. Avoid repeated freeze-thaw cycles.
1.1* Acid Stock Solution (125X):
Add 20 μL DMSO to *ate (component B) to make a 125X* acid stock solution.
2. Prepare standard solutions
*Salt standard solution
Add 50 μL of 1 mM KH2PO4 (Component C) to 950 μL of deionized water or enzyme reaction buffer to give a 50 μM * saline standard solution (PS7). A 50 μM * saline standard solution (PS7) was taken and serially diluted 1:2 to obtain a serially diluted phosphate standard with deionized water or enzyme reaction buffer.
3. Prepare a working solution
Add 20 μL of 125X* stock solution to 2.5 mL of sterile H2O and mix well to make a working solution of *salt. Avoid potential Pi contamination. Note: Avoid direct exposure of *salts (component B) to light. Due to the high sensitivity of this assay to Pi, it is extremely important to use Pi-free labware and reagents.
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文献和实验【求助】关于构建目的基因启动子序列的虫荧光素酶报告基因重组质粒的问题请教
应该有现成的商品化质粒可买吧,犯不上自己构建吧,你可以上网查查或者问一下公司。 longgyin8341 太好了,谢谢楼上的,碧云天公司有! 另外,不知道谁能提供雌激素受体(estrogen receptor,ER)的带荧光素酶的报告重组质粒?或者能告诉我那些地方可以买到?万分感谢! wangleisdnu Why you do not do Real time PCR? you could test
一家小公司的产品,不过实验室一直用,效果一直有保证,同期进行连接的其他同学也都连接成功。 结果: 转化后,平板上均有数量不等的菌落产生,少时4-5个,多时10来个。摇菌扩增8 h,进行菌液PCR检测(含阳性对照),检测组无阳性结果,但一直在1000 bp左右出现较弱条带,阳性组正常。提取质粒后,10 μl体系双酶切2 h,竟然在2000 bp和3000 bp处出现较弱条带(比Marker稍暗)。 特此悬赏10个叮当,请高手指点迷津。
由于各家公司生产的热循环仪提供的各种实验方案不太一致,所以优化的策略也不尽相同,然而在各种方案中,前文所述的那些影响实时PCR 反应的因素却总是相通的。只要能较好地控制实验条件,优化实验步骤,要想获得满意的实验结果并不一定都是困难的。下面本文将就影响到实验结果的一些基本参数和实验步骤谈一谈关于实时定量PCR的优化。 1. 基本参数的优化: 1.1 MgCl2的浓度 在PCR反应中,MgCl2的浓度对影响酶的活性是至关重要的,不仅如此,合适的MgCl
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